The invention relates to improved chromatography medium
Conventional chromatography media is based on spherical particles which can be either porous (mesoporous) or nonporous. These particles are normally packed in a column body in such a way as to produce maximum particle density. The particle density for such columns should be high in order to avoid the presence of random voids within the column bed which exhibit a detrimental effect on both chromatographic efficiency and physical stability of the packed column. Columns produced in this manner provide good chromatographic performance and good physical stability. From a theoretical point of view, chromatographic media packed in this manner approximate a dense assembly of spheres arranged in a cubic close pack geometry. Assuming the spheres are of uniform diameter, columns packed in a cubic close pack geometry will exhibit excellent physical stability and nearly theoretical chromatographic performance.
One problem with this packing geometry is that it imposes serious constraints on operating pressure when using high-performance stationary phases. Chromatographic efficiency is inversely proportional to particle diameter so in order to achieve improved chromatographic performance, this can only be achieved by using small diameter particles. Pressure for a column packed with spheres in a cubic close pack geometry is related to the operating flow rate, column diameter, column length, operating temperature, mobile phase viscosity and particle diameter. Column pressure is inversely proportional to the square of the particle diameter. For that reason, reduction in particle size when using conventional packed column based on particulate chromatographic media results in a geometric increasing pressure while yielding only a linear increase in chromatographic performance. For example, reducing the particle diameter by a factor of two should theoretically double the chromatographic performance of the column but the column pressure will increase by a factor of four.
The history of evolution of chromatographic materials in modern HPLC and ion chromatography has included a progressive decrease in the particle diameter in order to provide improved performance chromatographic materials. Now, however, the particle diameter has reached the point where significant improvements require dramatic increases in equipment costs along with significant reductions in instrument reliability and ease-of-use in order to deal with further pressure increases.
Furthermore, use of small particle size chromatographic media is often combined with reduction in column length in order to minimize the associated increase in pressure. Unfortunately, chromatographic performance is directly proportional to column length. Accordingly, improvements in chromatographic performance had been minimal as improved chromatographic efficiency in terms of plates per meter when using smaller diameter particle size chromatographic media is largely offset by reductions in column length resulting in similar chromatographic performance in terms of plates per column. While short columns with small diameter particle size chromatographic media make possible rapid separations for applications where high resolution is not a critical requirement, many chromatographic separations require improvements in terms of chromatographic performance. This can only be achieved by increasing column length or by reducing particle diameter while maintaining constant column length.
One technology which has been employed in order to address the above problems is the preparation of monolithic materials. A monolithic material is a chromatographic material composed of a continuous chromatographic medium with a series of through pores providing a means of fluid flow through the porous structure. Such a structure provides a significant potential advantage over particulate chromatographic media because it is theoretically possible to independently control the size of the through pores and the volume fraction of the stationary phase. Such materials have been prepared in a wide variety of shapes and sizes from a wide variety of materials. Surface area of such materials is controlled to the introduction of secondary mesopores in addition to the macropores which provide the fluid flow path through the monolithic material. Because the control of the pore size is independent of the effective size of the stationary phase segments, porosity and column pressure can be controlled independently of chromatographic performance.
There are several significant issues with this technology. First, preparation of materials of suitable chromatographic performance, operating pressure and physical stability require considerable experimentation. Furthermore, optimization is highly material dependent and so relatively minor changes in media composition require reoptimization of the entire preparation protocol. For this reason, development of new chromatographic media represents a major development project. This contrasts substantially with the situation with conventional particulate chromatographic media where a given particle platform can easily be modified to provide a wide variety of chromatographic media with minimal development effort.
An even more serious drawback to monolithic chromatographic media is the problem of shrinkage. Preparation of monolithic materials generally involves filling a cylindrical housing with the materials necessary for preparation of the monolith. After allowing the monolith to form (either through polymerization of monomers or condensation of inorganic precursors) a cylindrical porous monolithic rod is produced. The problem associated with this preparation process is that the resulting rod is smaller then the cylindrical housing in which it was produced. This presents a serious obstacle to the preparation of high performance chromatographic media. If the monolith is left in the housing in which it was initially prepared, fluid will preferentially flow around the monolith rather than flowing through it since the pressure drop for fluid flowing in the gap between the monolith and the cylindrical housing is significantly lower than the pressure drop for fluid flow through the monolith.
Mixed-bed columns packed with a bed comprising a mixture of strong anion and cation exchange resin have been used for liquid chromatography. The literature teaches that the cation and anion exchange resins form agglomerates or clumps which are undesirable because of poor flow distribution of liquid through the bed, e.g., channeling, which is taught to lead to low utilization of the resin's ion exchange capacity and a generally inefficient ion exchange operation. Various techniques have been suggested to avoid the perceived undesirable agglomeration. (U.S. Pat. Nos. 4,347,328 and 3,168,485 and European Patent Application EP1,241,083 A1.)
There is a need to overcome the weaknesses of both conventional particulate chromatographic media and conventional monolithic materials.